Project Summary How neural circuits are assembled during development remains a central unsolved problem in the neurosciences. Finding the genes that control the specificity of synaptic connections within circuits will be one of the keys to eventually manipulating circuit components and promoting functional regeneration following injury. However, elucidating circuit connectivity has been challenging for a number of reasons, including the complexity of the circuits themselves, issues of functional redundancy and the scarcity of genetic tools in many systems to manipulate the development and activity of defined neuronal subsets. This project uses a novel approach that capitalizes on the advanced genetics of Drosophila to define a neural circuit sufficient to drive locomotor behavior, but simple enough that we can determine the relevant synaptic connections and identify the genes controlling its development. Using a temperature-sensitive mutation, shibirets1 that conditionally blocks synaptic transmission at elevated temperatures, neurons within the relatively simple larval nervous system will be synaptically inactivated. By adding back synaptic activity to selected subsets of neurons by targeted expression of wild-type shibire+, a minimal circuit of interneurons, together with sensory and motor neurons, capable of generating motor output will be identified. The long- term goal of this project is to develop Drosophila larval locomotion as a model system for studying circuit development. Given the conservation of nervous system development and function within the animal kingdom, many of the rules by which neural circuits in Drosophila are synaptically connected during development are likely to be directly applicable to understanding the development of mammalian circuits.